Cell-free synthesis of Herpes simplex virus (HSV) proteins has been achieved by using a messenger dependent lysate (MDL) derived from rabbit reticulocytes. Many viral proteins ranging in size from greater than or equal to 130,000 daltons to less than 20,000 daltons have been synthesized in a cell-free system. In addition to cell-free experiments, in vivo pulse-labeled proteins from HSV infected cells, using both defective and nondefective forms of HSV, have been studied on long polyacrylamide gels and characterized as to whether they are early or late genomic products of HSV infection. Certain polypeptides in cells infected with defective HSV have been shown to bind to DNA cellulose. One of these binding proteins is low molecular weight (approximately 20,000 daltons) and appears to be similar to a low molecular weight protein synthesized in the MDL cell-free system using RNA derived from defective HSV-infected cells.